The effect of nucleoside on Na reabsorption via Na /nucleoside cotransporter in cultured rat epididymal epithelia was studied by short-circuit current (Isc) technique. Guanosine added apically stimulated Isc in a dose-dependent manner, with a median effective concentration (EC50) of 7 ± 2 μM (mean ± SEM). Removal of Na from the apical bathing solution or pretreatment with a nonspecific Na /nucleoside cotransporter inhibitor, phloridzin, completely blocked the Isc response to guanosine. Moreover, the guanosine response was abolished by pretreatment of the tissue with ouabain, a Na /K -ATPase inhibitor, suggesting the involvement of Na /nucleoside cotransporter on the apical side and Na /K -ATPase on the basolateral side in Na reabsorption. In contrast, the Isc response to guanosine was not affected after desensitization of purinoceptors by ATP. Addition of the Na /K /2Cl− symport inhibitor bumetanide to the basolateral side or the nonspecific Cl− channel blocker diphenylamine-2-carboxylate to the apical side showed no effect on the Isc response to guanosine, excluding stimulation of Cl− secretion by guanosine as the cause of the guanosine-induced Isc. The Isc response to purine nucleoside (guanosine and inosine) was much higher than that to pyrimidine nucleoside (thymidine and cytidine). Consistent with substrate specificity, results of reverse transcription-polymerase chain reaction revealed mRNA for concentrative nucleoside transporter (CNT2), which is a purine nucleoside-selective Na /nucleoside cotransporter in the epididymis, but not for CNT1. It is suggested that the Na /nucleoside cotransporter (i.e., CNT2) may be one of the elements involved in Na and fluid reabsorption in the epididymis, thereby providing an optimal microenvironment for the maturation and storage of spermatozoa.
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